Separation of intermediates of iron-catalyzed dopamine oxidation reactions using reversed-phase ion-pairing chromatography coupled in tandem with UV-visible and ESI-MS detections.

Reversed-phase ion-pairing chromatography (RP-IPC) is coupled on-line with electrospray ionization-mass spectrometry (ESI-MS) through an interface comprising a four-way switch valve and an anion exchange column. Regeneration of the anion exchange column can be accomplished on-line by switching the four-way switch valve to interconnect the column to a regeneration solution. Positioning the anion exchange column between the RP-IPC and ESI-MS instruments allows the ion-pairing reagent (IPR) sodium octane sulfonate to be removed. The IPC-ESI-MS method enabled us to separate and detect four intermediates of the Fe(III)-catalyzed dopamine oxidation. In particular, 6-hydroxydopamine, which is short-lived and highly neurotoxic, was detected and quantified. Together with the separation of other intermediates, gaining insight into the mechanism and kinetics of the Fe(III)-catalyzed dopamine oxidation becomes possible.